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Background: Rodents are recognized as the hosts of many vector-borne bacteria and protozoan parasites and play an important role in their transmission and maintenance. Intensive studies have focused on their infections in vectors, especially in ticks, however, vector-borne bacterial and protozoan infections in rodents are poorly understood although human cases presenting with fever may due to their infection have been found. Methods: From May to October 2019, 192 wild rodents were trapped in wild environment of Guangxi Province, and the spleen samples were collected to reveal the presence of vector-borne bacterial and protozoan infections in them. The microorganisms in rodents were identified by detecting their DNA using (semi-)nested PCR. All the PCR products of the expected size were subjected to sequencing, and then analyzed by BLASTn. Furthermore, all the recovered sequences were subjected to nucleotide identity and phylogenetic analyses. Results: As a result, 192 rodents representing seven species were captured, and Bandicota indica were the dominant species, followed by Rattus andamanensis. Based on the (semi-)nested PCR, our results suggested that Anaplasma bovis, Anaplasma capra, Anaplasma ovis, Anaplasma phagocytophilum, "Candidatus Neoehrlichia mikurensis", "Candidatus E. hainanensis", "Candidatus E. zunyiensis", three uncultured Ehrlichia spp., Bartonella coopersplainsensis, Bartonella tribocorum, Bartonella rattimassiliensis, Bartonella silvatica, two uncultured Bartonella spp., Babesia microti and diverse Hepatozoon were identified in six rodent species. More importantly, six species (including two Anaplasma, two Bartonella, "Ca. N. mikurensis" and Bab. microti) are zoonotic pathogens except Anaplasma bovis and Anaplasma ovis with zoonotic potential. Furthermore, dual infection was observed between different microorganisms, and the most common type of co-infection is between "Ca. N. mikurensis" and other microorganisms. Additionally, potential novel Bartonella species and Hepatozoon species demonstrated the presence of more diverse rodent-associated Bartonella and Hepatozoon. Conclusions: The results in this work indicated great genetic diversity of vector-borne infections in wild rodents, and highlighted the potential risk of human pathogens transmitted from rodents to humans through vectors. Author summary: Rodents are the important hosts of many pathogens, especially vector-borne pathogens, and can harbor and transmit them to humans and cause diseases. In China, little information is known about the infection of vector-borne pathogens in rodents although the rodent species are very diverse. In this study, seven wild rodent species were collected to identify the genetic diversity of vector-borne bacteria and protozoan. Interestingly, diverse vector-borne microorganisms, including four Anaplasma, five Ehrlichia, "Candidatus Neoehrlichia mikurensis", six Bartonella, Babesia microti and diverse Hepatozoon were identified in six rodent species. Furthermore, two potential novel Bartonella species were detected. All these data suggested great genetic diversity of vector-borne pathogens infections in rodents. Of them, six species (including two Anaplasma, two Bartonella, "Ca. N. mikurensis" and Bab. microti) are pathogenic to humans except A. bovis and A. ovis with zoonotic potential, suggesting significant risks to local human population. [ABSTRACT FROM AUTHOR]

Plague, one of the most devastating infectious diseases in human history, is caused by the bacterium Yersinia pestis. Since the 1950s, the Dehong Dai–Jingpo Autonomous Prefecture (DH) in Yunnan Province, China, has recorded plague outbreaks that have resulted in 1,153 human cases and 379 deaths. The genetic diversity and transmission characteristics of Y. pestis strains in this region remain unknown. Here, we performed high-resolution genomic epidemiological analysis of 175 Y. pestis strains isolated from five counties and 19 towns in DH between 1953 and 2007. Phylogenetic analysis revealed that most DH strains were located in lineage 1.ORI2, which could be further subdivided into seven sub-phylogroups (SPG1-SPG7). The dominant sub-phylogroups of Y. pestis in DH varied during different periods and presented a population shift. Genomic evidence showed that plague might have emerged from the southwest of DH (e.g., Longchuan or Ruili counties) or its bordering countries, and subsequently spread to the northeast in multiple waves between 1982 and 2007. Our study infers a fine-scale phylogeny and spread pattern of the DH Y. pestis population, which extends our knowledge regarding its genetic diversity and provides clues for the future prevention and control of plague in this region. Author summary: Yersinia pestis, the causative agent of plague, has led to three historic pandemics and remains a major public health issue. Here, genomic epidemiological analysis of 175 Y. pestis strains isolated over 50 years from the Dehong Dai—Jingpo Autonomous Prefecture, a region located in southwestern China, revealed the genomic evolution scenario of Y. pestis with high resolution. Our results revealed that plague epidemics in this region were led by different phylogroups, and population shift events were observed during different years, suggesting high dynamics of Y. pestis populations in the local natural plague foci. We found that the spread of plague was from southwest to northeast and inferred the possible origin of the epidemics, which would help strengthen surveillance of the epizootic in local natural plague foci and promote the prevention and control of human plague. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the occurrence and molecular characteristics of Bartonella infections in small rodents in the Shangdang Basin, China. Small rodents were captured using snap traps, and their liver, spleen, and kidney tissues were harvested for Bartonella detection and identification using a combination of real-time PCR of the ssrA gene (296 bp) and conventional PCR and sequencing of the gltA gene (379 bp). Results showed that 55 of 147 small rodents to be positive for Bartonella, with a positivity rate of 37.41%, and 95% confidence interval of 29.50%- 45.33%. While the positivity rate across genders (42.62% in males and 33.72% in females, χ2 = 1.208, P = 0.272) and tissues (28.57% in liver, 33.59% in spleen, and 36.76% in kidney, χ2 = 2.197, P = 0.333) of small rodents was not statistically different, that in different habitats (5.13% in villages, 84.44% in forests, and 54.17% in farmlands, χ2 = 80.105, P<0.001) was statistically different. There were 42 Bartonella sequences identified in six species, including 30 B. grahamii, three B. phoceensis, two B. japonica, two B. queenslandensis, one B. fuyuanensis and four unknown Bartonella species from Niviventer confucianus, Apodemus agrarius and Tscherskia triton. In addition to habitat, Bartonella species infection could be affected by the rodent species as well. Among the Bartonella species detected in this area, B. grahamii was the dominant epidemic species (accounting for 71.43%). B. grahamii exhibited four distinct clusters, and showed a certain host specificity. In addition, 11 haplotypes of B. grahamii were observed using DNASP 6.12.03, among which nine haplotypes were novel. Overall, high occurrence and genetic diversity of Bartonella were observed among small rodents in the Shangdang Basin; this information could potentially help the prevention and control of rodent-Bartonella species in this area. Author summary: Bartonella species are recognized as emerging zoonotic pathogens that are mainly transmitted by blood-sucking arthropods, some being infectious to humans. Rodents are important natural reservoirs of Bartonella species. Bartonella infection has been detected in various countries worldwide, and their prevalence in rodents varies greatly across countries and regions. This is the first report of Bartonella prevalence and genetic diversity in Shangdang Basin of China. It revealed a high Bartonella-positivity rate in Niviventer confucianus and Apodemus agrarius in this area. Relatively higher occurrence of Bartonella species was detected in forests and farmlands. Six Bartonella species were detected, B. grahamii, B. phoceensis, B. japonica, B. queenslandensis, B. fuyuanensis and one unknown Bartonella species, of which B. grahamii was the dominant species, with high genetic diversity, and potentially pathogenicity in humans. Our study provided a better understanding of the occurrence and genetic diversity of Bartonella species in the small rodents in Shangdang Basin. [ABSTRACT FROM AUTHOR]

Cryptococcus gattii (C. gattii) is a fungal pathogen that once caused an outbreak of cryptococcosis on Vancouver Island, and had spread worldwide, while few data were available in China. In this study, seven clinical isolates of C. gattii VGII were collected from 19 hospitals, Multi-locus Sequence Typing (MLST) analysis and whole-genome sequencing (WGS) was performed, combined with published data for phylogenetic analysis. In addition, in vitro antifungal susceptibility testing, phenotypic analysis, and in vivo virulence studies were performed, subsequently, histopathological analysis of lung tissue was performed. C.gattii VGII infected patients were mainly immunocompetent male, and most of them had symptoms of central nervous system (CNS) involvement. MLST results showed that isolates from China exhibited high genetic diversity, and sequence type (ST) 7 was the major ST among the isolates. Some clinical isolates showed a close phylogenetic relationship with strains from Australia and South America. All clinical isolates did not show resistance to antifungal drugs. In addition, there was no correlation between virulence factors (temperature, melanin production, and capsule size) and virulence while in vivo experiments showed significant differences in virulence among strains. Lung fungal burden and damage to lung tissue correlated with virulence, and degree of damage to lung tissue in mice may highlight differences in virulence. Our work seeks to provide useful data for molecular epidemiology, antifungal susceptibility, and virulence differences of C. gattii VGII in China. Author summary: Cryptococcus gattii is one of the life-threatening fungal pathogens that can infect immunocompetent individuals. C.gattii can be invaded through the respiratory tract and can be spread to the brain, causing meningoencephalitis and even death. In 1999, an outbreak of C. gattii VGII occurred on Vancouver Island in Canada and spread in the Pacific Northwest region, which attracted more attention. Compared with other nations, the data of C.gattii VGII in China was limited. To better understand the molecular epidemiology and microbiological characteristics of C.gattii VGII isolates from China, bioinformatics analysis and in vivo and in vitro experiments were performed for clinical strains. Multi-locus Sequence Typing and whole-genome sequencing reveal the genetic diversity of the isolates and genetic relationships with isolates from other countries. The antifungal drug susceptibility and virulence differences of the isolates were also investigated. The authors expect that our work can provide useful data for the prevention and control of C. gattii VGII in China and to draw more attention to C. gattii. [ABSTRACT FROM AUTHOR]

Background: Leptospirosis is one of the most important neglected tropical infectious diseases worldwide. Icterohaemorrhagiae has been throughout recent history, and still is, the predominant serogroup of this pathogen in China. However, very little in detail is known about the serovars or genotypes of this serogroup. Methodology/Principal Findings: In this study, 120 epidemic strains from five geographically diverse regions in China collected over a 50 year period (1958~2008), and 8 international reference strains characterized by 16S rRNA sequencing and MLST analysis. 115, 11 and 2 strains were identified as L. interrogans, L. borgpetersenii, and L. kirschneri, respectively. 17 different STs were identified including 69 ST1 strains, 18 ST17, 18 ST128, 9 ST143 and 2 ST209. The remaining 12 strains belonged to 12 different STs. eBURST analysis demonstrated that, among the clonal complexes isolated (CCs), CC1 accounted for 73.3% (88/120) strains representing three STs: ST1, ST128 and ST98. ST1 was the most likely ancestral strain of this CC, followed by singleton CC17 (17/120) and CC143 (11/120). Further analysis of adding 116 serogroup Icterohaemorrhagiae strains in the MLST database and studies previously described using global eBURST analysis and MST dendrogram revealed relatively similar ST clustering patterns with five main CCs and 8 singletons among these 244 strains. CC17 was found to be the most prevalent clone of pathogenic Leptospira circulating worldwide. This is the first time, to our knowledge, that ST1 and ST17 strains were distributed among 4 distinct serovars, indicating a highly complicated relationship between serovars and STs. Conclusions/Significance: Our studies demonstrated a high level of genetic diversity in the serogroup Icterohaemorrhagiae strains. Distinct from ST17 or ST37 circulating elsewhere, ST1 included in CC1, has over the past 50 years or so, proven to be the most prevalent ST of pathogenic leptospires isolated in China. Moreover, the complicated relationship between STs and serovars indicates an urgent need to develop an improved scheme for Leptospira serotyping. Author Summary: Leptospirosis, caused by pathogenic Leptospira spp, is a globally widespread zoonosis. In this study, our focusing on serogroup Icterohaemorrhagiae strains of Leptospira using MLST as a tool for phylogenetic analysis that has led to a better understanding of evolution of Leptospira. This totally consisted of 120 epidemic strains from five geographically diverse regions were isolated over the past 50 years in China and 8 strains from seven different countries. 17 STs were identified in these 128 strains by MLST analysis. Adding 116 serogroup Icterohaemorrhagiae in the Leptospira MLST database and studies previously described, 22 STs were identified in the 244 isolates. The genetic diversity of Leptospira belonging to serogroup Icterohaemorrhagiae from China was generally different from that of isolates elsewhere. Results of the 16S rRNA sequencing typing and MLST genotyping method were nearly consistent. Here, MLST revealed the high diversity of STs among the serogroup Icterohaemorrhagiae strains in China. Our present study provides a blueprint for further phylogenetic research. More convenient molecular techniques have to be developed to identify and characterize Leptospira species and STs. [ABSTRACT FROM AUTHOR]

Background: Schistosoma japonicum still causes severe parasitic disease in mainland China, but mainly in areas along the Yangtze River. However, the genetic diversity in populations of S. japonicum has not been well understood across its geographical distribution, and such data may provide insights into the epidemiology and possible control strategies for schistosomiasis. Methodology/Principal Findings: In this study infected Oncomelania snails were collected from areas in the middle and lower (ML) reaches of the Yangtze River, including Hubei, Hunan, Anhui, Jiangxi and Jiangsu provinces, and in the upper reaches of the river, including Sichuan and Yunnan provinces in southwest (SW) China. The adult parasites obtained from experimentally infected mice using isolated cercariae were sequenced individually for several fragments of mitochondrial regions, including Cytb-ND4L-ND4, 16S-12S and ND1. Populations in the ML reaches exhibited a relatively high level of diversity in nucleotides and haplotypes, whereas a low level was observed for populations in the SW, using either each single fragment or the combined sequence of the three fragments. Pairwise analyses of F-statistics (Fst) revealed a significant genetic difference between populations in the ML reaches and those in the SW, with limited gene flow and no shared haplotypes in between. It is rather obvious that genetic diversity in the populations of S. japonicum was significantly correlated with the geographical distance, and the geographical separation/isolation was considered to be the major factor accounting for the observed difference between populations in the ML reaches and those in the SW in China. Conclusions: S. japonicum in mainland China exhibits a high degree of genetic diversity, with a similar pattern of genetic diversity as observed in the intermediate host snails in the same region in China. Author Summary: Despite the existing threat of schistosomiasis in some rural areas along the Yangtze River, the genetic diversity of Schistosoma japonicum has not been investigated across its wide geographical distribution in China, and such information may provide insight into the disease epidemiology and the development of its control measures. In this study, the adult parasites, obtained through infecting mice with cercariae from snails of the genus Oncomelania collected from a wide range of localities in currently endemic areas of schistosomiasis in the middle and lower (ML) reaches of the Yangtze River, and in Sichuan and Yunnan provinces in the upper reaches of the river in southwest (SW) China, were sequenced individually for mitochondrial genes. In general, a relatively high degree of genetic variation was observed in populations in the ML reaches in terms of nucleotide and haplotype diversity, but a low level was observed in populations in the SW. The significant difference in genetic diversity as revealed by F-statistics, and the existence of no shared haplotypes, were observed between populations in the ML reaches and those in the SW, indicating the effect of geographical separation/isolation upon the schistosomes and probably the parasite-snail system in China. [ABSTRACT FROM AUTHOR]

Background: Oncomelania hupensis is the unique intermediate host of Schistosoma japonicum, which causes schistosomiasis endemic in the Far East, and especially in mainland China. O. hupensis largely determines the parasite's geographical range. How O. hupensis's genetic diversity is distributed geographically in mainland China has never been well examined with DNA sequence data. Methodology/Principal Findings: In this study we investigate the genetic variation among O. hupensis from different geographical origins using the combined complete internal transcribed spacer 1 (ITS1) and ITS2 regions of nuclear ribosomal DNA. 165 O. hupensis isolates were obtained in 29 localities from 7 provinces across mainland China: lake/marshland and hill regions in Anhui, Hubei, Hunan, Jiangxi and Jiangsu provinces, located along the middle and lower reaches of Yangtze River, and mountainous regions in Sichuan and Yunnan provinces. Phylogenetic and haplotype network analyses showed distinct genetic diversity and no shared haplotypes between populations from lake/marshland regions of the middle and lower reaches of the Yangtze River and populations from mountainous regions of Sichuan and Yunnan provinces. The genetic distance between these two groups is up to 0.81 based on Fst, and branch time was estimated as 2–6 Ma. As revealed in the phylogenetic tree, snails from Sichuan and Yunnan provinces were also clustered separately. Geographical separation appears to be an important factor accounting for the diversification of the two groups of O. hupensis in mainland China, and probably for the separate clades between snails from Sichuan and Yunnan provinces. In lake/marshland and hill regions along the middle and lower reaches of the Yangtze River, three clades were identified in the phylogenetic tree, but without any obvious clustering of snails from different provinces. Conclusions: O. hupensis in mainland China may have considerable genetic diversity, and a more complex population structure than expected. It will be of significant importance to consider the genetic diversity of O. hupensis when assessing co-evolutionary interactions with S. japonicum. Author Summary: The intermediate host of Schistosoma japonicum in Asia is the snail Oncomelania hupensis, which can be separated phenotypically into ribbed- and smooth-shelled morphotypes. In China, the typical morphotype is ribbed-shelled, with its distribution restricted to mainland China. Smooth-shelled snails with varix are also distributed in China, which are considered to belong to the same subspecies as the ribbed-shelled snails. In this study we investigate the genetic variation among O. hupensis from different geographical origins using combined complete ITS1 and ITS2 regions of nuclear ribosomal DNA. Snails including ribbed-shelled and smooth-shelled (but with varix on the shell) from the lake/marshland region of the middle and lower reaches of the Yangtze River, and smooth-shelled snails from mountainous regions of Sichuan and Yunnan provinces, were genetically distinct with no shared haplotypes detected. Furtheremore, the snails from Sichuan and Yunnan provinces were clustered in separate clades in the phylogenetic tree, and three clades were observed for snails from the middle and lower reaches of the Yangtze River. The population diversity of O. hupensis in China is thus considered large, and evolutionary relationships in the host-parasite system of O. hupensis-S. japonicum may be of interest for further research. [ABSTRACT FROM AUTHOR]